ABCMdb

ABCG2 p.Cys592Ser

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Publications

PMID: 16367905 [PubMed] Kage K et al: "Role of Cys-603 in dimer/oligomer formation of the breast cancer resistance protein BCRP/ABCG2."

No. Sentence Comment

62 Because BCRP protein expression levels in PA/C438S, PA/ C592S and PA/C608S cells were found to be remarkably decreased under non-reducing conditions (Fig. 2b), they were further examined by overexposing the immunoblot (Fig. 2c). Login to comment
69 For the PA /C592S and PA/C608S species, which may have mutations in the epitope for the 5D3 antibody located on a extracellular domain of BCRP, BCRP expression on the cell surface was undetectable by FACS, but protein could be detected at low levels by western blotting. Login to comment
71 Hence, the observed differences in BCRP expression, particularly in the PA/C438S, PA/C592S, PA/C603S and PA/C608S cells, are not attributable to low transcript levels. Login to comment
110 PA/C592S and PA/C608S showed decreased drug resistance and the sensitivity of PA/C438S cells was similar to that of the parental PA317 cells. Login to comment
122 Under non-reducing conditions, small quantities of both monomeric and dimeric forms of BCRP could be observed for the PA/C438S, PA/C592S and PA/C608S transfectants as well as in PA/C603S cell extracts following overexposure of the blot (Fig. 2c). Login to comment

PMID: 18644784 [PubMed] Ozvegy-Laczka C et al: "Interaction with the 5D3 monoclonal antibody is regulated by intramolecular rearrangements but not by covalent dimer formation of the human ABCG2 multidrug transporter."

No. Sentence Comment

254 They found that C592S and C608S had impaired 5D3 binding; however, these two mutants showed very low expression levels in this study (9). Login to comment

PMID: 20518788 [PubMed] Shigeta J et al: "BCRP/ABCG2 confers anticancer drug resistance without covalent dimerization."

No. Sentence Comment

29 In the present study, to elucidate the possible involvement of Cys-592 and Cys-608 in BCRP dimerization and protein activity, we established double (C592SÆC603S, C592SÆC608S, and C603SÆC608S) and triple (C592SÆC603SÆC608S) mutant BCRP transfectants, and compared the expression, dimer formation, and function of these BCRP mutants with wild-type and single (C592S, C603S, and C608S) mutant proteins. Login to comment
33 The wild-type and single (C592S, C603S, and C608S) mutant BCRP cDNAs in the same vector were also used as controls. Login to comment
63 PA317 cells were transfected with the wild-type, single (C592S, C603S, and C608S), double (C592SÆC603S, C592SÆC608S, and C603SÆC608S), and triple (C592SÆC603SÆC608S) mutant BCRP cDNAs in pHa-IRES-DHFR bicistronic expression vector as described previously. Login to comment
67 The resulting mixed populations of the drug-selected cells were designated as PA /WT-mix, PA / C592S-mix, PA /C603S-mix, PA /C608S-mix, PA /C592SÆC603S-mix, PA/ C592SÆC608S-mix, PA /C603SÆC608S-mix, and PA /C592SÆC603SÆC608S-mix. Login to comment
77 Breast cancer resistance protein (BCRP) dimer formation was observed in PA /WT-mix, PA /C592S-mix, PA /C592SÆC603S-mix, and PA/C592SÆC608S-mix cells. Login to comment
82 The BCRP expression levels and monomer /dimer formation in PA/ WT and single mutant BCRP transfectants (PA /C592S, PA/C603S, and PA/C608S) were essentially the same as those in our previous report (Fig. 3a-d). Login to comment
95 The ratio of 75-kDa BCRP in PA/C592S, PA /C592SÆC608S, and PA/C592SÆC603SÆC608S cells was two-third to half of the total BCRP. Login to comment
97 +SH -SH PA/WT-mix PA/C592S-mix PA/C603S-mix PA/C608S-mix PA/C592S·C603S-mix PA/C592S·C608S-mix PA/C603S·C608S-mix PA/C592S·C603S·C608S-mix PA317 PA/WT-mix PA/WT-mix(0.5) PA/C592S-mix PA/C603S-mix PA/C608S-mix PA/C592S·C603S-mix PA/C592S·C608S-mix PA/C603S·C608S-mix PA/C592S·C603S·C608S-mix PA317 150 75 kDa 150 75 kDa GAPDH (b)(a) Fig. 2. Login to comment
102 -mix -cl.1 -cl.8 PA317 PA/WT PA/C592S PA/C603S PA/WT-mix -mix -cl.3 -cl.7 PA317 PA/WT-mix -mix -cl.5 -cl.7 PA317 PA/C608S PA/WT-mix -mix -cl.6 -cl.7 PA317 BCRP (-SH) 75 150 kDa GAPDH (+SH) BCRP (+SH) PA/WT-mix -mix -cl.1 -cl.4 PA317 PA/WT-mix -mix -cl.6 -cl.7 PA317 PA/WT-mix -mix -cl.6 -cl.7 PA317 PA/C592S ·C603S PA/C592S ·C608S PA/C603S ·C608S PA/C592S ·C603S·C608S PA/WT-mix -mix -cl.2 -cl.7 PA317 BCRP (-SH) 75 150 kDa GAPDH (+SH) BCRP (+SH) (a) (b) (c) (d) (e) (g)(f) (h) Fig. 3. Login to comment
105 (a) PA / WT cells; (b) PA / C592S cells; (c) PA / C603S cells; (d) PA / C608S cells; (e) PA / C592SÆC603S cells; (f) PA / C592SÆC608S cells; (g) PA / C603SÆC608S cells; (h) PA / C592SÆC603SÆ C608S cells. Login to comment
110 PA /C592S transfectants exhibited intermediate levels of resistance to SN-38 whereas PA /C608S transfectants showed only marginal levels of resistance to this drug. Login to comment
120 Based on the results of drug sensitivity assay, we selected PA /WT-mix, PA/C592S-cl.6, PA /C603S-cl.3, PA /C592SÆC608S-cl.7, and PA/C592SÆC603SÆC608S-cl.7 cells as representative drug-resistant transfectants for further study. Login to comment
122 As shown in Figure 7(a), intracellular mitoxantrone was at low levels in PA /WT-mix and PA /C603S-cl.3 cells and at intermediate levels in PA /C592S-cl.6, PA /C592SÆC608S-cl.7, and PA /C592SÆC603SÆC608S-cl.7 cells. Login to comment
134 Immunofluorescent microscopy analysis using an anti-BCRP antibody BXP-21 was performed to examine whether 250 200 70-kDa BCRP 75-kDa BCRP 100 150 0 50 NormalizedBCRPexpression(%ofPA/WT) PA317 -mix -cl.1 -cl.8 -mix -cl.6 -cl.7 -mix -cl.3 -cl.7 -mix -cl.5 -cl.7 -mix -cl.1 -cl.4 -mix -cl.2 -cl.7 -mix -cl.6 -cl.7 -mix -cl.6 -cl.7 PAC592S ·C603S PA/C592S ·C608S PA/C603S ·C608S PA/C592S ·C603S·C608S PA/WT PA/C592S PA/C603S PA/C608S Fig. 4. Login to comment
138 SN-38 resistance of mutant BCRP transfectants Cell line IC50 to SN-38† (nM) Degree of resistance‡ (x-fold) Relative expression level Total BCRP§ 75-kDa BCRP- PA317 3.7 ± 0.2 1 - - PA / WT-mix 90 ± 4.4 24 100 100 PA / WT-cl.1 172 ± 6.1 47 207 206 PA / WT-cl.8 77 ± 10.3 21 56 56 PA / C592S-mix 12.4 ± 0.8 3.4 34 24 PA / C592S-cl.6 28.5 ± 1.4 7.7 52 37 PA / C592S-cl.7 13.7 ± 0.5 3.7 16 11 PA / C603S-mix 9.7 ± 0.5 2.6 54 51 PA / C603S-cl.3 409 ± 15.9 110 146 132 PA / C603S-cl.7 158 ± 17.7 43 135 122 PA / C608S-mix 4.8 ± 0.3 1.3 27 3 PA / C608S-cl.5 5.6 ± 0.2 1.5 26 7 PA / C608S-cl.7 6.3 ± 0.6 1.7 43 12 PA / C592SÆC603S-mix 3.6 ± 0.2 1.0 27 10 PA / C592SÆC603S-cl.1 5.1 ± 1.1 1.4 27 10 PA / C592SÆC603S-cl.4 6.2 ± 0.8 1.7 43 26 PA / C592SÆC608S-mix 16.7 ± 1.8 4.6 92 67 PA / C592SÆC608S-cl.2 22.8 ± 2.0 6.2 163 111 PA / C592SÆC608S-cl.7 34.5 ± 4.9 9.4 151 106 PA / C603SÆC608S-mix 2.9 ± 0.2 0.8 21 3 PA / C603SÆC608S-cl.6 4.4 ± 0.4 1.2 57 3 PA / C603SÆC608S-cl.7 3.4 ± 0.1 0.9 51 4 PA / C592SÆC603SÆC608S-mix 4.1 ± 0.2 1.1 13 7 PA / C592SÆC603SÆC608S-cl.6 4.0 ± 0.4 1.1 25 13 PA / C592SÆC603SÆC608S-cl.7 11.1 ± 1.5 3.0 145 102 †The cells were incubated for 4 days at 37°C in the presence or absence of various concentrations of 7-ethyl-10-hydroxycamptothecin (SN-38). Login to comment
150 As shown in Table 2, FTC (1 lM) completely reversed the resistance of PA /WT-mix and PA /C592S-cl.6 cells to SN-38 and mitoxantrone. Login to comment
163 (16) PA /C608S cells also expressed a BCRP monomer whereas PA/ C592S cells expressed a BCRP dimer. Login to comment
172 Introduction of the C592S mutation into BCRP-C603S extinguishes the counter partner against Cys-608 in the intramolecular disulfide bridge. Login to comment
195 PA/C592S-cl.6 4. Login to comment
208 (1) PA317 cells; (2) PA / WT-mix cells; (3) PA / C592S-cl.6 cells; (4) PA / C603S-cl.3 cells; (5) PA / C592SÆC608S-cl.7 cells; (6) PA / C592SÆC603SÆC608S-cl.7 cells. Login to comment
226 PA/WT, PA /C592S, PA/C603S, PA /C592SÆC608S, and PA /C592SÆC603SÆC608S-cl.7 cells exhibited drug resistance and low accumulation of mitoxantrone. Login to comment
228 Significant expression of a 75-kDa BCRP was also found in PA /C592S, PA /C592SÆC608S, and PA/ C592SÆC603SÆC608S-cl.7 cells, whereas the PA/C608S cells mainly expressed a 70-kDa BCRP PA317 PA/WT-mix PA/C592S·C603S ·C608S-cl.7 BCRP Nuclei Merge 20 µm 20 µm 20 µm 20 µm 20 µm 20 µm 20 µm 20 µm 20 µm Fig. 8. Login to comment
234 Effect of FTC on the SN-38 and mitoxantrone resistance of mutant BCRP transfectants Cell line SN-38 Mitoxantrone Relative expression level of total BCRP§ Degree of resistance† RI‡ Degree of resistance† RI‡ )FTC +FTC )FTC + FTC PA317 1.0 ± 0.1 0.9 ± 0.1 - 1.0 ± 0.1 0.9 ± 0.1 - - PA / WT-mix 19.3 ± 1.7 1.4 ± 0.1 13.8 9.0 ± 0.3 1.2 ± 0.1 7.5 100 PA / C592S-cl.6 4.2 ± 0.3 1.2 ± 0.1 3.5 3.7 ± 0.1 1.3 ± 0.1 2.8 52 PA / C603S-cl.3 106 ± 7.3 7.9 ± 0.3 13.4 57 ± 5.9 15 ± 0.7 3.8 146 PA / C603S-cl.7 18 ± 2.0 4.1 ± 0.2 4.4 28 ± 3.9 12 ± 1.0 2.3 135 PA / C592SÆC608S-cl.7 8.4 ± 0.5 4.0 ± 0.2 2.1 3.3 ± 0.2 3.3 ± 0.1 1.0 151 PA / C592SÆC603SÆC608S-cl.7 4.1 ± 0.3 4.6 ± 0.3 0.9 7.5 ± 0.2 6.0 ± 0.8 1.3 145 †The cells were incubated for 4 days at 37°C in the presence or absence of various concentrations of anticancer agents in the presence or absence of 1 lM fumitremorgin C (FTC). Login to comment
245 Fumitremorgin C (FTC) strongly reversed the resistance of PA /WT and PA /C592S cells to SN-38 and mitoxantrone, and partially reversed the drug resistance of PA /C603S cells. Login to comment

PMID: 20812902 [PubMed] Ni Z et al: "Structure and function of the human breast cancer resistance protein (BCRP/ABCG2)."

No. Sentence Comment

136 Importantly, Shigeta et al. showed that the triple-mutant C592S/C603S/C608S retained substantial resistance to mitoxantrone, topotecan, and SN-38 [86]. Login to comment
300 The recent study by Shigeta et al. [86] also demonstrated that the triple-mutant C592S/C603S/C608S in PA317 cells retained substantial activity with some alteration in substrate recognition, although its level of expression was significantly decreased and its subcellular distribution was altered. Login to comment

PMID: 25036722 [PubMed] Szafraniec MJ et al: "Determinants of the activity and substrate recognition of breast cancer resistance protein (ABCG2)."

No. Sentence Comment

209 Position Type of mutation Effect on the transporter References NBD Lys 86 Met (i) No stimulation of the ATPase activity by prazosin; (ii) no influence on the transport of mitoxantrone Henriksen et al. (2005b) Glu 126 stop, Phe 208 Ser, Ser 248 Phe, Glu 334 stop Inability to transport hematoporphyrin Tamura et al. (2006) Glu 211 Gln Complete abolishment of the ATPase activity and methotrexate transport Hou et al. (2009) Pro 392 Ala Significant reduction in the efflux activity of mitoxantrone, BODIPY-prazosin and Hoechst 33342 Ni et al. (2011) TM1 Gly 406 Ala Gly 410 Ala No influence on the activity of the transporter Polgar et al. (2004) Gly 406 Leu Gly 410 Leu (i) Loss of the ability to transport rhodamine123; (ii) impaired transport of mitoxantrone, Pheide and BODIPY-prazosin Polgar et al. (2004) Extracellular loop 1 Phe 431 Leu (i) Loss of the ability to transport methotrexate; (ii) 10% level of hematoporphyrin transport compared to the WT protein Tamura et al. (2006) Ser 441 Asn Inability to transport hematoporphyrin Tamura et al. (2006) Ser 441 Asn Loss of the ability to transport methotrexate Tamura et al. (2006) TM2 Lys 452 Ala His 457 Ala Increase in transport of mitoxantrone, BODIPY-prazosin and Hoechst 33342 Cai et al. (2010) Lys 453 Ala Arg 465 Ala Decrease in transport of mitoxantrone, BODIPY-prazosin, Hoechst 33342, doxorubicin, SN-38 and rhodamine 123 Cai et al. (2010) TM3 Arg 482 Gly Arg 482 Thr (i) No change in the inhibitory activity of lapatinib; (ii) about two times greater inhibition by ritonavir, saquinavir and nalfinavir than in the WT variant; (iii) gaining the ability to transport rhodamine123 and doxorubicin; (iv) no influence on the transport of mitoxantrone; (v) loss of the ability to transport methotrexate Dai et al. (2008), Gupta et al. (2004), Honjo et al. (2001), Mitomo et al. (2003) Arg 482 Thr (i) Lower IC 50 of cyclosporine A for mutant than for WT variant; (ii) lower elacridar inhibition potency Xia et al. (2007) Arg 482 Lys Complete loss of transport activity Ejendal et al. (2006) Phe 489 Leu Impaired transport of porphyrins, no transport of methotrexate Tamura et al. (2006) Extracellular loop 3 Asn 590 Tyr Over twice reduced transport of mitoxantrone, topotecan, daunorubicin and rhodamine 123 Vethanayagam et al. (2005) Cys 592 Ala/Cys 608 Ala (i) Transport of mitoxantrone almost unchanged; (ii) transport of BODIPY-prazosin significantly impaired Henriksen et al. (2005a) Extracellular loop 3 Cys 603 Ser Cys 592 Ser/Cys 608 Ser Cys 592 Ser/Cys 603 Ser/Cys 608 Ser Diminished susceptibility to the inhibitory activity of fumitremorgin C Shigeta et al. (2010) Cys-less Arg 482 Gly-BCRP Complete loss of the ability to efflux mitoxantrone Liu et al. (2008b) The positions of the amino acid residues refer to the topological model of BCRP proposed by Wang et al. (2009). Login to comment
230 Cys592 Ser/Cys603 Ser-BCRP and Cys592 Ser/Cys608 Ser-BCRP were detected as dimers, while Cys603 Ser/Cys608 Ser-BCRP and Cys592 Ser/Cys603 Ser/Cys608 Ser-BCRP as monomers, suggesting that no other Cys are involved in dimerization. Login to comment
231 At the same time, cells transfected with a triple mutant, Cys592 Ser/Cys603 Ser/Cys608 Ser-BCRP, showed significant drug resistance and a low accumulation of mitoxantrone which indicated that dimer formation might not be required for BCRP functioning as a transporter. Login to comment